How cancer develops is now understood. As described by Robert Weinberg, “How Cancer Arises”, Scientific American, September 1996, the normal, healthy human body has about 30 trillion cells which live in a complex interdependent body. The nucleus of each cell includes chromosomes having DNA molecules that contain genes. Each gene specifies a sequence of amino acids that must be linked together to make a particular protein; this protein then carries out the work of the gene. When a gene is switched on, the cell responds by synthesizing the encoded protein. Mutations in a gene can perturb a cell by changing the amounts or activities of the protein product.
Normal cells replicate only when certain biochemical conditions, internal and external to the cell, are met. The cells of a tumor are now known to descend from a single cell that begins a program of uncontrolled reproduction because of an accumulation of successive mutations, usually occurring over a long time, in specific classes of genes within the cell.
Two gene classes are instrumental in triggering cancer. Proto-oncogenes encourage cell growth, and tumor suppressor genes inhibit it. When mutated, proto-oncogenes can drive excessive cell multiplication. If, in addition, the tumor suppressor genes are mutated or turned off by other cell abnormalities, the cell loses the ability to prevent runaway growth.
Recently, biomedical science has determined that stimulatory and inhibitory pathways in a cell are carefully regulated by a complex set of processes that occur in during a cell cycle. In a normal cell, the cycle integrates the mixture of growth-regulating signals and decides whether the cell should pass through a life cycle as set forth in FIG. 1.
The life cycle is composed of four stages: In the G1 (gap 1) phase, the cell increases in size and prepares to copy its DNA. This copying occurs in the following S (synthesis) stage which enables the cell to duplicate its complement of chromosomes. After synthesis, a second gap period, G2, occurs during which the cell prepares for mitosis (M), the time when the enlarged parent cell divides in half to produce two daughter cells, each with a complete set of chromosomes. The new daughter cells immediately enter G1 and may go through the complete cycle or enter the G0 phase where cycling stops.
The difficulty in detecting cancer arises from the subtle onset of the disease in a single cell embedded in a host organ comprising billions of cells. Pathologists routinely rely on microscopic examination of cell morphology using methods that originated over a hundred years ago. These staining methods are labor-intensive, time-consuming, and frequently in error. New micro-analytical methods for high speed (real time) automated screening of tissues and cells are critical to advancing pathology and hold the potential for improving diagnosis and treatment of cancer patients.
Novel technologies to assess these diseases in their early development are crucial to effect a successful treatment and recovery. Promising techniques under investigation include non-invasive but expensive magnetic resonance imaging, inexpensive ultrasound imagining of organs or organ linings, or minimally invasive sampling of minute body tissues or fluids in microdevices. The resolution of MRI imaging is currently in the range of hundreds of microns and limited by the strength of the magnetic field and its gradient. Ultrasound resolution is limited by the wavelength of sound, and is similarly limited to hundreds of microns. Optical and Laser-based spectroscopies and imaging are inherently higher resolution, being limited only by the optical diffraction of light to hundreds of nanometers. Nanoscopic tools like the near-field optical scanning microscope push the resolution to tens of nanometers (the size of protein molecule) and are limited by fiber optic tip microfabrication techniques.
Another technique that holds promise for detection of cellular changes is the laser biocavity as disclosed in the aforementioned U.S. patent application Ser. No. 09/489,247 and related patent application Ser. No. 09/221,331, both by Paul L. Gourley, and U.S. Pat. No. 5,793,485 by Paul L. Gourley et al. The disclosure of each of these references is incorporated herein by reference thereto.
As shown in FIG. 2, the invention described in the aforementioned references incorporates a glass chip with a semiconductor laser, the chip having input and output reservoirs (on either sides of the figure) to store a small quantity of material to be tested, and a plurality of channels having a cross-sectional area on the same order of magnitude as a blood cell. A vacuum is applied to the output reservoir to pull material through the channels where it forms part of the optical path of the laser, as shown, enabling high throughput analyses of flowing biofluids.
The semiconductor nanolaser is the enabling component of this microanalysis system because of its ability to emit coherent, intense light from a small aperture compatible with the dimensions of a human cell. By permitting fluid flow through surface-emitting semiconductor geometry, there is provided a means for high throughput screening of cells, particulates and fluid analytes in a sensitive microdevice. As these fluids flow through these channels, their components will alter the emitted lasing spectrum. The spectral shifts in the lasing frequencies can then be used to measure changes in the cells.
Most importantly, cells can be analyzed in their physiologic condition as removed from the body. There are no time delays or difficulties associated with tagging cells with stains or fluorescent markers. Thus, the usual time delay of tissue pathology under the microscope is eliminated. And, the lasing technique has been found to detect subtle changes in cellular compositions that are orders of magnitude smaller than can be observed by standard optical microscopy. In addition, this device does not alter or manipulate the cellular components, thus allowing for direct observations of nuclear and cellular events.
The applications of such a portable biological sensing device are far-reaching. One potential surgical application of this biocavity laser is a “smart scalpel,” for online flow cytometry in the operating room. Conventional flow cytometers tend to be large, expensive instruments that require highly trained personnel for tagging the cells and operating the instrument. A portable cytometer system designed for an operating room setting could analyze minute volumes of cells and improve the success rate of tumor resection. Such a system could decrease costs and improve patient survival rates by avoiding tumor regrowth and subsequent surgeries. To achieve this capability it is necessary to understand the basic operation of the laser and its ability to detect biologically relevant events.